Protection of the world's natural resources must be a priority to ensure global human and animal health. Aquatic environments are essential natural resources that must be maintained, monitored and studied. Humans and wildlife rely on aquatic environments for a range of services. In addition, freshwater environments are relatively scarce across the landscape (<0.08% of the world's surface) and these ecosystems are integral points of interaction among organisms. Infectious diseases in freshwater environments are a major concern for medical, veterinary and conservation efforts world-wide. Many pathogens of humans and wildlife depend on these freshwater ecosystems. However, our understanding of many freshwater pathogens is limited. Therefore, it is imperative that we learn more about the interactions taking place within aquatic environments to ensure that these natural resources will be available and in good health for our future generations. Ranaviruses (RVs) (family Iridoviridae, genus Ranavirus) are large, double-stranded DNA viruses that infect economically and ecologically important freshwater poikilothermic vertebrates including fish, amphibians and reptiles. RVs are a unique group of viruses for two reasons: (i) No other group of viruses infect such a wide diversity of hosts (i.e. amphibians, fish and reptiles), except perhaps for the orthomyxoviruses (e.g. influenza viruses); (ii) Members of the genus Ranavirus have undergone multiple evolutionarily recent hosts species shifts, jumping from fish to amphibians, amphibian species to amphibian species, amphibians to reptiles and perhaps from amphibians and/or reptiles to fish. In spite of the global impact RVs have on economically and ecologically important hosts, our current understanding of the mechanisms of RV pathogenesis, including the genetic determinants that contribute to RV pathogenesis and host range, is extremely limited. To address this issue, we will test the hypothesis that 7 RV genes are involved with viral pathogenesis. Testing this hypothesis will include innovative in vitro and in vivo assays, including gene knock down by siRNA and gene knock out by homologous recombination before assaying pathogenesis in the virus' natural host, tiger salamanders. The results obtained will be a direct result of undergraduate and Master degree student's research efforts and the outcome of these experiments will be significant contributions to our understanding of the evolution of dsDNA virus host-pathogen interactions, and a more comprehensive understanding of the molecular determinants of RV pathogenesis. In addition, the data obtained from this application will be instrumental in maintaining our aquatic natural resources for future generations. PUBLIC HEALTH RELEVANCE: Protection of the world's natural resources must be a priority to ensure global human and animal health. Freshwater environments are relatively scarce across the landscape and infectious diseases in freshwater environments are a major concern for medical, veterinary and conservation efforts world-wide. Using ranaviruses as a model system, this application will explore the host-pathogen interactions of these important pathogens of lower vertebrates and will contribute to our understanding of emerging infectious diseases of freshwater environments, shed light into the evolution of dsDNA virus pathogenic determinants and give insight into the evolution of the host response to viral infections.